RPB2 Gene Research Articles

Additions to Taiwan Fungal Flora 1: Neomassariaceae fam. nov.

In the course of surveys on dothideomycetous fungal genera associated with various hosts in Taiwan, Neomassaria-like species were collected as saprobes on dead stems of Rhododendron sp. Maximum likelihood and Bayesian phylogenetic analyses based on concatenated LSU, rpb2, SSU and tef1 gene matrices indicate that Neomassaria-like isolates generated in our study formed a separate clade in a sister group relationship with Neomassaria fabacearum with high statistical support. Hence, the new species Neomassaria formosana is described to accommodate the new linage in Neomassaria. The new species is characterised by immersed to erumpent, papillate ascomata with a dark brown peridium, fissitunicate, cylindrical to oblong asci and fusoid to ellipsoid, hyaline, bicellular ascospores with 4–6 large guttules. Moreover, Neomassaria formed a monophyletic and well-supported lineage with an uncertain phylogenetic placement within the Pleosporales. Therefore, the new family Neomassariaceae is proposed for the genus Neomassaria within the order Pleosporales, Dothideomycetes. The morphological characters also support justification for both the new family and the new species.

A Ready-to-Use Single- and Duplex-TaqMan-qPCR Assay to Detect and Quantify the Biocontrol Agents Trichoderma asperellum and Trichoderma gamsii.

Trichoderma asperellum strain icc012 and Trichoderma gamsii strain icc080, the microbial active ingredients of RemedierTM (ISAGRO, Novara, Italy), are biocontrol agents (BCAs) employable for crop protection against a wide range of fungal pathogens, including soil-borne pathogens and fungi involved in grapevine trunk disease. In this study, single and duplex real-time quantitative PCR (qPCR) methods to detect and quantify T. asperellum and T. gamsii were developed. Primers/probe sets were designed on the T. asperellum and T. gamsii rpb2 genes and tested for specificity on a panel of microorganisms commonly associated with grape wood and soil. No differences were observed comparing single- and duplex-qPCR assays on different BCAs, 1 pg of target DNA was detected approximately at Cq = 34. R2-values and the efficiency were always equal to 0.99 and >80%, respectively. The detection limit of the duplex-qPCR assay on artificially inoculated samples was 2 × 103 and 4 × 104 conidia g-1 of grape wood tissue and soil, respectively. The methods will be useful to better schedule BCA application in the field and in grapevine nurseries, as well as for investigating the dynamic of BCA populations.

First Report ofAlternaria alternataCausing Fruit Rot of Grapes in Pakistan

In July 2016, a severe fruit rot was observed on table grapes (Vitis vinifera) from five different fruit packing houses in the Rawalpindi district (33°38′19.2″N, 73°01′45.0″E) of Punjab province, Pakistan. During the survey, dark gray to black mycelium was observed on the surface of the berries and rapidly expanded to entire bunches. After 3 to 5 days of postharvest storage at ambient temperature, prevalence of fruit rot was 100% in all locations of Rawalpindi district. A total of 200 symptomatic berries were surface disinfested with 1% sodium hypochlorite for 2 min, rinsed three times with sterile distilled water, and then dried on a filter. Excised pieces of sterile tissue (4 mm²) at lesion margins were transferred to potato dextrose agar (PDA) and incubated at 25 ± 2°C for 3 to 5 days. Fungal cultures derived from the isolates gave rise to colonies that were dark olivaceous green with light brown margins, velvety, appressed, and cottony at the center. Conidiophores were septate, bent or sometimes branched, 25.6 to 55.4 μm long and 2.5 to 3.6 μm wide. Conidia were borne singly or in short chains and were obpyriform to obclavate, measuring 9.5 to 55.5 × 6.1 to 25.6 μm with zero to three longitudinal and two to six transverse septa. Morphological characters matched those of Alternaria alternata (Fr.) Keissl (Simmons 2007). For molecular identification, ITS1, 5.8S; ITS2, RNA polymerase II (RPB2); and endopolygalacturonase (endoPG) gene of two representative isolates (Alt 01 and Alt 05) were amplified with primers ITS1/ITS4 (White et al. 1990), fRPB2-7cF/fRpb2-7cR (Liu et al. 1999), and PG3/PG2b (Andrew et al. 2009), respectively. The amplicons were sequenced and deposited in GenBank (accession nos. MG763130, MF785102, MG773211, MG773212, MG786192, and MG786193). A BLAST search of the GenBank database revealed close identity (98 to 100%) with the sequences of various A. alternata isolates (accession nos. KU182490 and MF141012 for ITS gene, KX938348 and KX938349 for RPB2 gene, and KY969535 and KY969535 for endoPG gene) of A. alternata in NCBI. To confirm pathogenicity, 10-µl aliquots of conidial suspension (10⁶ spores/ml) were pipetted onto three nonwounded and four wounded asymptomatic grape berries per isolate. The berries were incubated at 25 ± 2°C in sterile moist chambers, and the experiment was conducted twice. After 3 days, dark gray to black mycelium, similar in appearance to the original isolates, developed on both wounded and nonwounded inoculated berries, whereas control berries inoculated with sterile distilled water remained symptomless. The morphology of the fungus isolated on PDA was identical to that of the original cultures. A. alternata has been reported to cause postharvest fruit rot on grapes in South Africa (Swart and Holz 1991) and Slovakia (Kakalikova et al. 2009). To our knowledge, this is the first report of A. alternata causing fruit rot of grapes in Pakistan, where the disease poses a significant threat to sustainability of grape producers.

Roof-Inhabiting Cousins of Rock-Inhabiting Fungi: Novel Melanized Microcolonial Fungal Species from Photocatalytically Reactive Subaerial Surfaces.

Subaerial biofilms (SAB) are an important factor in weathering, biofouling, and biodeterioration of bare rocks, building materials, and solar panel surfaces. The realm of SAB is continually widened by modern materials, and the settlers on these exposed solid surfaces always include melanized, stress-tolerant microcolonial ascomycetes. After their first discovery on desert rock surfaces, these melanized chaetothyrialean and dothidealean ascomycetes have been found on Mediterranean monuments after biocidal treatments, Antarctic rocks and solar panels. New man-made modifications of surfaces (e.g., treatment with biocides or photocatalytically active layers) accommodate the exceptional stress-tolerance of microcolonial fungi and thus further select for this well-protected ecological group. Melanized fungal strains were isolated from a microbial community that developed on highly photocatalytic roof tiles after a long-term environmental exposure in a maritime-influenced region in northwestern Germany. Four of the isolated strains are described here as a novel species, Constantinomyces oldenburgensis, based on multilocus ITS, LSU, RPB2 gene phylogeny. Their closest relative is a still-unnamed rock-inhabiting strain TRN431, here described as C. patonensis. Both species cluster in Capnodiales, among typical melanized microcolonial rock fungi from different stress habitats, including Antarctica. These novel strains flourish in hostile conditions of highly oxidizing material surfaces, and shall be used in reference procedures in material testing.

Gray mold disease of Paris polyphylla var. yunnanensis and its pathogen identification

Gray mold disease is one of the most important diseases of planted Paris polyphylla var. yunnanensis, the disease appeared primarily as blossom blights and fruit rots, but also as stem rots, leaf rots.In this study, the pathogenetic fungi was isolated from plant tissue or sclerotia that covering the fruit of diseased P. polyphylla var. yunnanensis, the pathogen was certified according to Koch's Postulation. The pathogen produced abundant black, irregular sclerotia on surface of diseased plants and potato dextrose agar. The conidiophores and clusters of oval conidia resembled a grape-like cluster, the size of conidia was 9.70-13.70 μm [average of (11.32±0.82)μm]×7.05-9.12 μm [average of (8.24±0.48)μm], the microconidia produced on potato dextrose agar were spherical,and the size was （3.34±0.31） μm,the pathogen was identified as Botrytis sp based on morphological characteristics. The DNA sequence analysis of the G3PDH, HSP60, RPB2 genes placed the pathogen in a single clade that outside defined species of Botrytis, so the pathogen could be identified as a new species of Botrytis. The pathogen requires 20 °C, pH 8, darkness or low light condition for the best growth.

Collapsimycopappus: A new leaf pathogen with mycopappus‐like propagules in Mycosphaerellaceae

AbstractA previously undescribed mycopappus‐like fungus associated with large zonate leaf spot disease of Styrax obassia was found in Japan. Single‐propagule isolation of the fungus from necrotic areas of the diseased leaves yielded four pure cultures. Pathogenicity tests of the fungus were performed on potted S. obassia plants. Leaves were wounded with a sterilized needle and inoculated with mycelial agar. Development of necrotic zones was observed within 4 months. Numerous propagules appeared on the necrotic areas, and two strains of the same fungus were isolated again. The morphology of the pathogen was described, and its phylogeny was analysed based on the sequences of nuclear rDNA ITS, LSU regions and the rpb2 genes. Twenty‐four sequences were generated from six strains. Phylogenetic analyses indicated that the pathogen was closely related to the members of Mycosphaerellaceae, but was distinct in having brown propagules with hyphal appendages that collapsed at maturity and divided into part spores. Therefore, we propose Collapsimycopappus as a new genus to accommodate a single species, C. styracis.

Editorial Board

Lophiostoma bipolare was taxonomically revised based on the morphological observations and phylogenetic analyses of molecular data from nuclear rDNA SSU-ITS-LSU, TUB, tef1, and rpb2 genes. Twenty-nine strains were morphologically similar to Lo. bipolare. A total of 174 sequences were generated from the Lo. bipolare complex. Phylogenetic analyses based on TUB sequence revealed 11 distinct species within the Lo. bipolare complex. Morphological features of the ascospores and the anatomical structure of the ascomata from both field collections as well as axenic culture, which have been reported previously as variable features at intraspecific levels, were compared to evaluate the taxonomic reliability of these features. To clarify the generic position of the 11 species, phylogenetic analyses were done on SSU-ITS-LSU-tef1-rpb2 gene sequences. The Lo. bipolare complex shared phylogenetic relationships with Pseudolophiostoma and Vaginatispora, and formed an additional five distinct clades from other members of Lophiostomataceae. According to its phylogenetic position, Lo. bipolare sensu stricto was distantly related to Lophiostoma s. str., and formed an independent clade within Lophiostomataceae. Lophiostoma bipolare s. str. could be distinguished from the other lophiostomataceous genera by the clypeus around the ostiolar neck and by the thin and uniformly thick peridium. A novel genus described as Lentistoma was established to accommodate this species, and the epitypification of Lentistoma bipolare (basionym: Massarina bipolaris) was proposed. Other lineages of the Lo. bipolare complex could not be separated on the basis of the ascospore size and sheath variations, but were distinguished based on ascomatal features, such as the existence of the clypeus, brown hyphae surrounding the peridium, and the contexture of the peridium, which were stable indicators of generic boundaries in Lophiostomataceae. Four additional new genera with five new species were recognised based on these morphological differences: Crassiclypeus (C. aquaticus), Flabellascoma (F. cycadicola and F. minimum), Leptoparies (Lep. palmarum), and Pseudopaucispora (Pseudop. brunneospora). Three new species were added to Pseudolophiostoma (Pseudol. cornisporum, Pseudol. obtusisporum, and Pseudol. tropicum) and two new species were added to Vaginatispora (V. amygdali and V. scabrispora). The re-evaluation of the validity of several previously recognised genera resulted in the introduction of two new genera with new combinations for Lophiostoma pseudoarmatisporum as Parapaucispora pseudoarmatispora and Vaginatispora fuckelii as Neovaginatispora fuckelii.

Reporte del estado sexual de Lecanicillium longisporum en Melanaphis sacchari

Antecedentes: Durante la búsqueda de enemigos naturales de Melanaphis sacchari en Tecomán, Colima, México, se observaron poblaciones micosadas de pulgón amarillo en sorgo; tres meses después se encontraron estructuras fúngicas de reproducción sexual desarrollándose en la región dorsal abdominal de algunos pulgones.Objetivo: Dar a conocer por primera vez el estado teleomorfo de Lecanicillium longisporum, desarrollado sobre M. sacchari en Colima, México.Métodos: Se utilizaron técnicas de microscopía para el análisis morfológico de los peritecios encontrados sobre M. sacchari. Para la caracterización molecular se utilizó la región its y el gen rpb1. Resultados y conclusiones: Los peritecios encontrados en los pulgones momificados con L. longisporum son morfológicamente diferentes a Cordiceps confragosa y otras especies reportadas como teleomorfos de Lecanicillium. Las secuencias its y rpb1 mostraron 100% de similitud con las secuencias del anamorfo che-cnrcb 620 (its: kx426564; rpb1: kx426566) de L. longisporum. La identificación molecular confirmó que los peritecios corresponden al teleomorfo de la misma especie, siendo el primer reporte del estado sexual de L. longisporum.

Taxonomy and phylogeny of operculate discomycetes: Pezizomycetes

The class Pezizomycetes is monophyletic within the subdivision Pezizomycotina. The main distinguishing character of this class is operculate asci, although in some taxa this character has been lost. The circumscription of the families and generic level delimitation in Pezizomycetes is still controversial, although several molecular phylogenetic studies have been published on this group. This paper reviews 21 families of Pezizomycetes including five new families, which are introduced here, viz. Kallistoskyphaceae, Pseudombrophilaceae, Pulvinulaceae, Strobiloscyphaceae and Tarzettaceae. Moreover, this study provides a modified backbone tree based on phylogenetic analysis of five combined loci. Descriptions and illustrations of representative taxa for the families are provided from collections made in China, Thailand and the UK, herbarium material from international herbaria (FH, FLAS, H, HKAS and MA) and the literature. Pezizales separates into six major clades. Clade 1 of Pezizales includes the families Ascobolaceae and Pezizaceae. Clade 2 is the new family Kallistoskyphaceae. Clade 3 comprises the families Caloscyphaceae, Karstenellaceae and Rhizinaceae. Clade 4 represents the families Discinaceae, Helvellaceae, Morchellaceae, Tuberaceae and Underwoodia columnaris lineage. Clade 5 includes Chorioactidaceae, Sarcoscyphaceae and Sarcosomataceae and Clade 6 comprises Ascodesmidaceae, Glaziellaceae, Otideaceae, Pseudombrophilaceae, Pulvinulaceae, Pyronemataceae, Strobiloscyphaceae and Tarzettaceae. New sequence data belonging to ITS, LSU, SSU, TEF, RPB2 gene regions from 40 pezizalian species are provided here. The paper provides a working document for apothecial Pezizomycetes which can be modified as new data comes to light. It is hoped that by illustrating taxa we provide stimulation and interest in the operculate discomycetes, so that further research is carried out on this remarkable, but poorly studied group of fungi.

Determination of fungal pathogens of Hypera postica (Gyllenhall) (Coleoptera: Curculionidae): isolation, characterization, and susceptibility

BackgroundFungal pathogens of Hypera postica (Gyllenhall) (Coleoptera: Curculionidae) were collected from the vicinities of Adana and Igdir in Turkey. The pathogenicity of the fungal isolates against the pest were investigated. According to morphologic (colony morphology, spore shape) and molecular (sequences of ITS1-5.8S ITS2 region and EF1-α, Bloc, and RPB1 genes) characterizations, the isolates were identified as Beauveria bassiana (HpI-2, HpI-6, HpI-7, HpI-10, HpA-3, HpA-4, HpA-5) and Beauveria pseudobassiana (HpI-4). All these strains were isolated from H. postica for the first time. In order to determine pathogenesis of all isolates on the target pest, bioassays were conducted against larvae and adults, as screening of (1 × 107 conidia/ml) and dose-response (1 × 105, 1 × 106, 1 × 107, 1 × 108, conidia/ml), under laboratory conditions. The fungal isolates, closely related to each other, yielded significantly varied mortalities on larvae and adults. H. postica larvae were found more susceptible than adults to the fungal isolates in all tests. The highest mortality rates (100 and 98%) for larvae and adults, respectively, were obtained by B. bassiana strain HpA-5 within 14 days at 1 × 108 conidia/ml concentration. The median lethal concentration (LD50) of HpA-5 required to kill the larvae and adults of H. postica at concentrations of 2.37 × 104 and 1.4 × 105 conidia/ml, respectively. These results are promising; therefore, the B. bassiana strain HpA-5 can potentially be used against H. postica.

Phylogenetic position and taxonomy of Kusaghiporia usambarensis gen. et sp. nov. (Polyporales)

ABSTRACT A large polyporoid mushroom from the West Usambara Mountains in North-eastern Tanzania produces dark brown, up to 60-cm large fruiting bodies that at maturity may weigh more than 10 kg. It has a high rate of mycelial growth and regeneration and was found growing on both dry and green leaves of shrubs; attached to the base of living trees, and it was also observed to degrade dead snakes and insects accidentally coming into contact with it. Phylogenetic analyses based on individual and concatenated data sets of nrLSU, nrSSU and the RPB2 and TEF1 genes showed it, together with Laetiporus, Phaeolus, Pycnoporellus and Wolfiporia, to form a monophyletic group in Polyporales. Based on morphological features and molecular data, it is described as Kusaghiporia usambarensis.

Identification and Characterization of Alternaria iridiaustralis Causing Leaf Spot on Iris ensata in China

In 2016, a severe leaf spot disease was found on Iris ensata Thumb. in Nanjing, China. The symptom was elliptical, fusiform, or irregularly necrotic lesion surrounded by a yellow halo, from which a small-spored Alternaria species was isolated. The fungus was identified as Alternaria iridiaustralis based on morphological characteristics. The pathogenicity tests revealed that the fungus was the causal pathogen of the disease. Phylogenic analyses using sequences of ITS, gpd, endoPG, and RPB2 genes confirmed the morphological identification. This study is the first report of A. iridiaustralis causing leaf spots on I. ensata in China.

Multiple genes phylogeny and morphological characters reveal Dextrinoporus aquaticus gen. et sp. nov. (Polyporales, Basidiomycota) from southern China

Dextrinoporus aquaticus gen. et sp. nov. is proposed based on ITS, nLSU, RPB2, and mtSSU genes molecular phylogeny and morphological characteristics from southwestern China. The new species is characterized by an annual, pileate basidiocarp with a poroid hymenophore, a monomitic hyphal system, thick-walled generative hyphae with dextrinoid and cyanophilous reactions, presence of dendrohyphidia and ellipsoid, thin-walled, and acyanophilous basidiospores. Phylogenetic analyses based on four gene sequences indicate that D. aquaticus belongs to Polyporaceae in which it has an isolated position, and the new taxon is related to the clade of Lopharia cinerascens and Dentocorticium sulphurellum.

Resolving the Lophiostoma bipolare complex: Generic delimitations within Lophiostomataceae.

Lophiostoma bipolare was taxonomically revised based on the morphological observations and phylogenetic analyses of molecular data from nuclear rDNA SSU-ITS-LSU, TUB, tef1, and rpb2 genes. Twenty-nine strains were morphologically similar to Lo. bipolare. A total of 174 sequences were generated from the Lo. bipolare complex. Phylogenetic analyses based on TUB sequence revealed 11 distinct species within the Lo. bipolare complex. Morphological features of the ascospores and the anatomical structure of the ascomata from both field collections as well as axenic culture, which have been reported previously as variable features at intraspecific levels, were compared to evaluate the taxonomic reliability of these features. To clarify the generic position of the 11 species, phylogenetic analyses were done on SSU-ITS-LSU-tef1-rpb2 gene sequences. The Lo. bipolare complex shared phylogenetic relationships with Pseudolophiostoma and Vaginatispora, and formed an additional five distinct clades from other members of Lophiostomataceae. According to its phylogenetic position, Lo. bipolare sensu stricto was distantly related to Lophiostoma s. str., and formed an independent clade within Lophiostomataceae. Lophiostoma bipolare s. str. could be distinguished from the other lophiostomataceous genera by the clypeus around the ostiolar neck and by the thin and uniformly thick peridium. A novel genus described as Lentistoma was established to accommodate this species, and the epitypification of Lentistoma bipolare (basionym: Massarina bipolaris) was proposed. Other lineages of the Lo. bipolare complex could not be separated on the basis of the ascospore size and sheath variations, but were distinguished based on ascomatal features, such as the existence of the clypeus, brown hyphae surrounding the peridium, and the contexture of the peridium, which were stable indicators of generic boundaries in Lophiostomataceae. Four additional new genera with five new species were recognised based on these morphological differences: Crassiclypeus (C. aquaticus), Flabellascoma (F. cycadicola and F. minimum), Leptoparies (Lep. palmarum), and Pseudopaucispora (Pseudop. brunneospora). Three new species were added to Pseudolophiostoma (Pseudol. cornisporum, Pseudol. obtusisporum, and Pseudol. tropicum) and two new species were added to Vaginatispora (V. amygdali and V. scabrispora). The re-evaluation of the validity of several previously recognised genera resulted in the introduction of two new genera with new combinations for Lophiostoma pseudoarmatisporum as Parapaucispora pseudoarmatispora and Vaginatispora fuckelii as Neovaginatispora fuckelii.

Dominikia litorea, a new species in the Glomeromycotina, and biogeographic distribution of Dominikia

An arbuscular mycorrhizal fungus (AMF) producing clusters with colourless, small (11‒35 µm diam when globose) spores of unique morphological characters of two spore wall layers was grown in a trap culture and in single-species cultures. Both the spore wall layers are permanent and have the same thickness. The features of the spores prompted that the fungus most probably belongs to one of the genera, Dominikia or Kamienskia. Phylogenetic analyses of sequences of the SSU‒ITS‒LSU nrDNA and the RPB1 gene showed that the discussed AMF is an undescribed Dominikia sp. highly diverged molecularly from the 12 so far described species of the genus. Consequently, the fungus is described here as D. litorea sp. nov. The sporulation of D. litorea in the trap culture indicated that in the field the new species lived in mycorrhizal symbiosis with Xanthium spinosum that had colonized sand dunes of the Mediterranean Sea located near Verico, Greece. However, comparisons of the SSU‒ITS‒LSU sequences of D. litorea with those obtained from molecular environmental studies, which are deposited in public databases, indicated that the new species probably is also associated with roots of an unnamed plant species growing in China. In addition, based on available literature, sequence data and personal observations, the so far known geographical distribution, habitats, and plant-hosts of the described Dominikia spp. were presented and discussed. Finally, the potential participation of Dominikia spp. in influencing plants and plant communities with which they are associated and ecosystems in which they exist were discussed.

Families and genera of diaporthalean fungi associated with canker and dieback of tree hosts

In this study we accept 25 families in Diaporthales based on phylogenetic analyses using partial ITS, LSU, rpb2 and tef1-α gene sequences. Four different families associated with canker and dieback of tree hosts are morphologically treated and phylogenetically compared. These include three new families (Diaporthostomataceae, Pseudomelanconidaceae, Synnemasporellaceae), and one new genus, Dendrostoma (Erythrogloeaceae). Dendrostoma is newly described from Malus spectabilis, Osmanthus fragrans and Quercus acutissima having fusoid to cylindrical, bicellular ascospores, with three new species namely D. mali, D. osmanthi and D. quercinum. Diaporthostomataceae is characterised by conical and discrete perithecia with bicellular, fusoid ascospores on branches of Machilus leptophylla. Pseudomelanconidaceae is defined by conidiogenous cells with apical collarets and discreet annellations, and the inconspicuous hyaline conidial sheath when mature on Carya cathayensis, compared to morphologically similar families Melanconidaceae and Juglanconidaceae. Synnemasporellaceae is proposed to accommodate fungi with synnematous conidiomata, with descriptions of S. toxicodendri on Toxicodendron sylvestre and S. aculeans on Rhus copallina.

A new genus, Desertispora, and a new species, Diversispora sabulosa, in the family Diversisporaceae (order Diversisporales, subphylum Glomeromycotina)

Phylogenetic analyses of sequences of the SSU–ITS–LSU nrDNA segment and the RPB1 gene showed that the arbuscular mycorrhizal fungus originally described as Diversispora omaniana does not belong to the genus Diversispora, but represents a separate clade at the rank of genus in the family Diversisporaceae of the order Diversisporales. The closest natural relatives of the fungus proved to be species of the genera Corymbiglomus and Redeckera. Consequently, the new genus was named Desertispora, and Di. omaniana was renamed De. omaniana comb. nov. In addition, the morphological and histochemical features of spores and mycorrhizal structures of a new Diversispora sp., Di. sabulosa, were described and the closest relatives of the species were determined based on phylogenetic analyses of sequences of the two loci mentioned above. The new fungus was grown in single-species cultures established from spores extracted from a trap culture inoculated with a mixture of the rhizosphere soil and root fragments of Ammophila arenaria that had colonized maritime sand dunes of the Curonian Spit located in the north of Lithuania. Diversispora sabulosa was never found before in many different sites of the world which were sampled during the last 34 years by the last author of the paper. Also, the lack of molecular sequences in public databases of identity ≥ 97% to sequences of Di. sabulosa suggests that the fungus is rare on the Earth.

Ultrastructural and molecular characterization of Vairimorpha austropotamobii sp. nov. (Microsporidia: Burenellidae) and Thelohania contejeani (Microsporidia: Thelohaniidae), two parasites of the white-clawed crayfish, Austropotamobius pallipes complex (Decapoda: Astacidae).

The microsporidiosis of the endangered white-clawed crayfish Austropotamobius pallipes complex has generally been attributed to only one species, Thelohania contejeani, the agent of porcelain disease. Species identification was mostly assessed by macroscopic examination or microscopic evaluation of muscle samples rather than by molecular or ultrastructural analyses. A survey conducted on A. pallipes complex populations in Northern Italy highlighted the presence of two different microsporidia causing similar muscular lesions, T. contejeani and an undescribed octosporoblastic species Vairimorpha austropotamobii sp. nov. Mature spores and earlier developmental stages of V. austropotamobii sp. nov. were found within striated muscle cells of the thorax, abdomen, and appendages of the crayfish. Only octosporoblastic sporogony within sporophorous vesicles (SPVs) was observed. Diplokaryotic sporonts separated into two uninucleate daughter cells, which gave rise to a rosette-shaped plasmodium, and eight uninucleate spores were produced within the persistent SPV. Ultrastructural features of stages in the octosporoblastic sequence were similar to those described for Vairimorpha necatrix, the type species. Mature spores were pyriform in shape and an average of 3.9 × 2.2 µm in size. The polar filament was coiled 11-14 times, lateral to the posterior vacuole. The small subunit ribosomal RNA gene (SSU rRNA) and the large subunit RNA polymerase II gene (RPB1) of V. austropotamobii sp. nov. were sequenced and compared with other microsporidia. The highest sequence identity of SSU rRNA (99%) and RPB1 (74%) genes was with the amphipod parasite Nosema granulosis and subsequently with V. cheracis, which infects the Australian yabby Cherax destructor. In our work we discuss about the reasons for placing this new species in the genus Vairimorpha. In addition, we provide for T. contejeani a RPB1 gene sequence, supplemental sequences of SSU rRNA gene and ultrastructural details of its sporogony in the host A. pallipes complex.

Arthonia incarnata (Arthoniaceae), a rare and poorly known old‐growth forest lichen new to Asia

The rare and poorly known Northern Hemisphere old‐growth forest lichen Arthonia incarnata is reported for the first time for Asia from Japan and Korea. A detailed description and illustration is provided for the species based on the collections from Japan and South Korea. Bayesian and RAxML analyses of mtSSU, nLSU and RPB2 sequence data show A. incarnata to be only distantly related to the other Arthonia species hitherto sequenced and in particular to superficially similar species occurring in the same type of habitat such as A. didyma, A. spadicea and A. vinosa. For the analysis we generated new sequence data of A. incarnata for the mtSSU (5 specimens) and RPB2 (5) gene loci. Slight variation was observed in sequence data of the RPB2 gene between collections from eastern Asia and Sweden, but this is not substantiated by differences in morphology or ecological behaviour. Arthonia incarnata is confined to humid forests and only grows on parts of tree trunks shaded from precipitation and running water such as the lower side of leaning trunks or cavities at the base of trees. The species is found on trees with acidic bark, in eastern Asia mainly on Betula but also on coniferous trees and on dead wood. It was not found on bark of Salix (caprea), a common substrate for the species in Europe.

Outbreak of Fusarium oxysporum infections in children with cancer: an experience with 7 episodes of catheter-related fungemia

BackgroundFusarium species are widely spread in nature as plant pathogens but are also able to cause opportunistic fungal infections in humans. We report a cluster of Fusarium oxysporum bloodstream infections in a single pediatric cancer center.MethodsAll clinical and epidemiological data related to an outbreak involving seven cases of fungemia by Fusarium oxysporum during October 2013 and February 2014 were analysed. All cultured isolates (n = 14) were identified to species level by sequencing of the TEF1 and RPB2 genes. Genotyping of the outbreak isolates was performed by amplified fragment length polymorphism fingerprinting.ResultsIn a 5-month period 7 febrile pediatric cancer patients were diagnosed with catheter-related Fusarium oxysporum bloodstream infections. In a time span of 11 years, only 6 other infections due to Fusarium were documented and all were caused by a different species, Fusarium solani. None of the pediatric cancer patients had neutropenia at the time of diagnosis and all became febrile within two days after catheter manipulation in a specially designed room. Extensive environmental sampling in this room and the hospital did not gave a clue to the source. The outbreak was terminated after implementation of a multidisciplinary central line insertion care bundle. All Fusarium strains from blood and catheter tips were genetically related by amplified fragment length polymorphism fingerprinting. All patients survived the infection after prompt catheter removal and antifungal therapy.ConclusionA cluster with, genotypical identical, Fusarium oxysporum strains infecting 7 children with cancer, was most probably catheter-related. The environmental source was not discovered but strict infection control measures and catheter care terminated the outbreak.